: Resistance to endocrine therapy poses a serious challenge to the effective treatment of breast cancer. About one-third of all primary breast tumors lack estrogen receptor (ER) expression and are insensitive to endocrine therapy such as antiestrogens or ovarian ablation. This phenotype is associated with the worst prognosis. The mechanisms by which ER gene expression is abrogated are poorly understood. However, several recently reported studies suggest that epigenetic changes via DNA methylation may play a role in silencing expression of the ER gene as well as a variety of other tumor suppressor genes. Furthermore, the enzyme which catalyzes the methylation of CpG dinucleotides, DNA methyltransferase (DNA Mtase) is often overexpressed in cancer cells compared with normal cells, and its expression has been reported to increase with progression in some types of cancer. The studies proposed here address the overall hypothesis that loss of ER expression in some breast cancers is a consequence of loss of ER gene transcription associated with hypermethylation at a CpG island in the 5' regio of the human ER gene. They will further address the premise that increased DNA MTase expression may promote the progression of breast cancer to a hormone-independent phenotype by inhibiting expression of genes, like the ER, which would otherwise suppress that phenotype. The specific aims include: 1) to establish whether there is a relationship between increased expression of DNA MTase and loss of ER expression in human breast cancer cell lines and tissue specimens; 2) to examine the interaction between DNA methylation and histone acetylation as regulators of ER transcription and expression; and 3) to identify other CpG islands in addition to that of the ER that are consistently hypermethylated in hormone-independent human breast cancer cells since such CpG islands may identify genes that promote a hormone-sensitive phenotype. Taken together these studies should provide new insights into the biology of hormone resistance and possible novel therapeutic targets.